Staphylococci final

Dr Reena Kulshrestha, M.Sc, PhD

Dr Reena Kulshrestha, M.Sc, PhD*

ClassificationFamily

Genus

Species

MicrococcaceaeMicrococcus and StaphylococcusS. aureusS. saprophyticusS. epidermidisM. luteusmore than 20 speciesDr Reena Kulshrestha, M.Sc, PhD


INTRODUCTIONStaphyloccocci - derived from Greek stapyle (bunch of grapes)Gram positive cocci arranged in clustersHardy organisms surviving many non -physiologic conditionsInclude a major human pathogen and skin commensals



StreptococcusStaphylococcusDr Reena Kulshrestha, M.Sc, PhD


Grouping for Clinical Purposes1. Coagulase positive StaphylococciStaphylococcus aureus

2. Coagulase negative StaphylococciStaphylococcus epidermidisStaphylococcus saprophyticus



Coagulase-negative staphylococcus; frequently involved in nosocomial and opportunistic infections S. epidermidis lives on skin and mucous membranes; endocarditis, bacteremia, UTI S. hominis lives around apocrine sweat glands S. capitis live on scalp, face, external ear

All 3 may cause wound infections by penetrating through broken skin S. saprophyticus infrequently lives on skin, intestine, vagina; UTI



A. Staphylococcus aureusMajor human pathogen

Habitat - part of normal flora in some humans and animals

Source of organism - can be infected human host, carrier, fomite or environment



Grows in large, round, opaque colonies Optimum temperature of 37oC Facultative anaerobe Withstands high salt, extremes in pH, and high temperatures Carried in nasopharynx and skin Produces many virulence factors



Cultivation of S.aureusTemp. 10-42*C, pH 7.4 7.6Aerobes & facultative anaerobesNutrient Agar emulsifiable, smooth, shinyColony pigmentation - white, orange & yellow at 22*C, aerobic conditins, enhanced with 1% glycerol monoacetate or milkNA slope oil paint appearanceBlood Agar B hemolysisMac Conkeys Agar pink LF coloniesBroth media Salt- milk broth, Ludams medium uniform turbidity



Cell- associated virulence factorsCapsule or slime layer (glycocalyx) inhibits opsonisationPeptidoglycan (PG) rigidity to cell, activates complement, induces release of inflammatory cytokinesTeichoic acid is covalently linked to PG and is species specific:

- Facilitataes adhesion, protects from complement-mediated opsonisationDr Reena Kulshrestha, M.Sc, PhD


S. aureus- ribitol teichoic acid (polysaccharide A)S. epidermidis- glycerol teichoic acid (polysaccharide B)

Protein A - is covalently linked to PGchemotactic, antiphagocytic, anticomplementary, induces platelet damage & hypersensitivityBinds to Fc terminal of IgG



Clumping factor ( bound coagulase )Surface protein for Slide Coagulase test saline suspension of

S.aureus + Human plasma

Cocci are clumped

- Identification of S.aureus- Capsulated strain may not show



Virulence Factors Extracellular EnzymesCoagulase (free) - Antigenic, acts along with CRF Hyaluronidase spreading factor of S. aureus (Staphylokinase (fibrinolysin), fatty acid modifying enzymes & proteases ) Breaks down the connective tissueNucleaseCleaves DNA and RNA in S. aureusHeat stable



ProteaseStaphylokinase (fibrinolysin) facilitates adhesionLipases

- helps in infecting the skin and sub- cutaneous tissuesEsterases



Virulence Factors: ExotoxinsCytolytic (cytotoxins; cytolysins)Alpha toxin - hemolysin Reacts with RBCs, leucocidal, dermonecrotic, neurotoxic & lethalToxic to macrophages, lysosomes, muscle tissues, renal cortex & circulatory system Beta toxinSphingomyelinase, exhibits Hot-Cold phenomenonGamma toxinHemolytic activityDelta toxinCytopathic for:RBCs, Lymphocytes, Neutrophils, PlateletsEnterotoxic activity



Leucocidin- ( Panton- Valentine toxin)2 componenets S & F

*Synergohymenotropic toxins (leucocidin+gamma lysin)Enterotoxin- Causes Food poisoning Nausea, vomiting & diaarheaIP- 2-6 hrs, Heat stable- 100* for 10- 40 minsSource of infection food handler (carrier)Acts on autonomic nervous systemPotent- ug is toxigenicPyrogenic, mitogenic, hypotensive, thrombocytopenic & cytotoxic effectsDiagnosis Latex agglutination & ELISA



Exfoliative toxin (epidermolytic toxin) SSSS exfoliative skin diseases, effects new born Ritters disease -epidermis gets separated from the underlying tissues older patients - Toxic epidermal necrolysis Milder forms Pemphigus neonatorum & Bullous impetigo

Pyrogenic exotoxins- TSS fatal multisystem disease fever, hypotension, myalgia, vomiting, diarhhea, mucosal hyperemia & erythematous rashInfection of mucosal sites, skin & surgical woundsSuper Ags excessive & dis-regulated Immune Response -activates large no. of T- cells, IL-1 & 2, TNF, INF-y involves multisystem



Natural history of diseaseMany neonates, children, adults -intermittently colonized by S. aureusUsual sites - skin, nasopharynx, perineumBreach in mucosal barriers - can enter underlying tissueCharacteristic abscessesDisease due to toxin production



DISEASESDue to direct effect of organismLocal lesions of skinDeep abscessesSystemic infections

Toxin mediatedFood poisoningtoxic shock syndromeScalded skin syndrome



Factors predisposing to S. aureus infectionsHost factorsBreach in skinChemotaxis defectsOpsonisation defectsNeutrophil functional defectsDiabetes mellitusPresence of foreign bodies

Pathogen Factors Catalase (counteracts host defences)CoagulaseHyaluronidaseLipases (Imp. in disseminating infection)B lactasamase(ass. With antibiotic resistance)



PathogenesisPass skin first line of defense Benign infectionPhagocytosisAntibodyInflammatory responseChronic infectionsDelayed hypersensitivity



SKIN LESIONS

BoilsStyesFuruncles(infection of hair follicle)Carbuncles (infection of several hair follicles)Wound infections(progressive appearance of swelling and pain in a surgical wound after about 2 days from the surgery)Impetigo(skin lesion with blisters that break and become covered with crusting exudate)



DEEP ABSCESSSESCan be single or multipleBreast abscess can occur in 1-3% of nursing mothers in puerperiemCan produce mild to severe diseaseOther sites - kidney, brain from septic foci in blood



Systemic Infections

1. With obvious focusOsteomyelitis, septic arthritis2. No obvious focusheart (infective endocarditis)Brain(brain abscesses)3. Ass. With predisposing factors multiple abscesses, septicaemia(IV drug users)Staphylococcal pneumonia (Post viral)



B. TOXIN MEDIATED DISEASES1. Staphylococcal food poisoningDue to production of entero toxinsheat stable entero toxin acts on gutproduces severe vomiting following a very short incubation periodResolves on its own within about 24 hours



2. Toxic shock syndromeHigh fever, diarrhoea, shock and erythematous skin rash which desquamateMediated via toxic shock syndrome toxin10% mortality rateDescribed in two groups of patients Asso. with young women using tampones during menstruationDescribed in young children and men



3. Scalded skin syndromeDisease of young childrenMediated through minor Staphylococcal infection by epidermolytic toxin producing strainsMild erythema and blistering of skin followed by shedding of sheets of epidermisChildren are otherwise healthy and most eventually recover



Antibiotic sensitivity pattern

Very variable and not predictableVery imp. In Pt. ManagementMechanisms

1.B lactamase production - plasmid mediatedHas made S. aureus resistant to penicillin group of antibiotics - 90% of S. aureus (Gp A)B lactamase stable penicillins (cloxacillin, oxacillin, methicillin) used

2. Alteration of penicillin binding proteins(Chromosomal mediated)Has made S. aureus resistant to B lactamase stable penicillins10-20% S. aureus Gp (B) resistant to all Penicillins and Cephalasporins)Vancomycin is the drug of choice



Tested in lab using methicillinReferred to as methicillin resistant S. aureus (MRSA)Emerging problem in the worldIn Sri Lanka prevalence varies from 20- 40% in hospitalsDrug of choice - vancomycinIn Japan emergence of VIRSA(vancomycin intermediate resistant S. aureus)No effective antibiotics discovered -We might have to discover



DIAGNOSIS1. In all pus forming lesions Gram stain and culture of pus2. In all systemic infectionsBlood culture3. In infections of other tissuesCulture of relevant tissue or exudate



Identification of Staphylococcus in SamplesFrequently isolated from pus, tissue exudates, sputum, urine, and bloodCultivation, catalase, biochemical testing, coagulase



Mannitol Salts Agar (MSA)Staphylococcus aureusDr Reena Kulshrestha, M.Sc, PhD


Catalase2H2O2 O2 + 2H2OStreptococci vs. StaphylococciDifferential CharacteristicsDr Reena Kulshrestha, M.Sc, PhD


CoagulaseFibrinogen FibrinDifferential CharacteristicsDr Reena Kulshrestha, M.Sc, PhD


TreatmentDrain infected areaDeep/metastatic infections semi-synthetic penicllins cephalosporins erythromycin clindamycinEndocarditis semi-synthetic penicillin + an aminoglycoside



PreventionCarrier status prevents complete controlProper hygiene, segregation of carrier from highly susceptible individualsGood aseptic techniques when handling surgical instrumentsControl of nosocomial infections



2. Staphylococcus epidermidisSkin commensalHas predilection for plastic materialAss. With infection of IV lines, prosthetic heart valves, shuntsCauses urinary tract infection in cathetarised patientsHas variable ABS pattern Treatment should be aided with ABST



S. epidermidisLocationNormal skin flora opportunistic pathogenSkin/wound infectionsEndocarditisUTIExposureDirect contactNewbornsElderlyFomitesCathetersShuntsIV needlesProsthetics



S. saprophyticusPathogenesisFimbriaAdhesion proteinsAutolysinsDiseasesUTI/cystitisPeritonitisEnopthalmitisEndocaritisSeptic arthritis



S. xylosusCommensalIndustryFerment meatRed color of sausageFerment milkOrange color of cheesePathogenicityBiofilmsEnterotoxinsDiseaseNosocomialUTIFood poisoning (raw)



S. capitisEpidemiologySkin microbioticaHead predominantlyPathogenicityCoagulase (-)AB resistanceDiseasesValvular endocarditisNeonatal septicemiaOsteomyelitis



S. pseudointermediusAnimal microbioticaEpidemiologyZoonoticEnterotoxins+/- coagulaseDiseasesPyoderma (animals)Food poisoning



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Staphylococci final

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